Harris Gelbard - Academia.edu (original) (raw)

Papers by Harris Gelbard

Communicative & Integrative Biology, Nov 9, 2013

Virology, Dec 1, 1998

Simian immunodeficiency virus strain PBj14, SIV smmPBj14 , is unique among primate lentiviruses i... more Simian immunodeficiency virus strain PBj14, SIV smmPBj14 , is unique among primate lentiviruses in its ability to trigger the proliferation of resting simian lymphocytes and to cause the rapid death of experimentally inoculated pigtailed macaques. Severe enteropathy, immune activation, and extensive apoptosis, particularly within gut-associated lymphoid tissue, characterize the acute disease syndrome associated with SIV smmPBj14 infection. In the present study, we examined whether the ability of this virus to cause widespread apoptosis might be linked to the up-regulation of Fas ligand (CD95L) expression in virally infected cells. In vitro studies revealed that expression of the viral Nef protein, in the absence of any other viral gene product, was sufficient to up-regulate the transcriptional activity of the CD95L promoter and to cause cell surface expression of Fas ligand. This up-regulation was NFAT dependent (inhibited by cyclosporin A) and did not occur in cells that expressed a mutated derivative of the viral Nef protein, lacking a previously defined immunoreceptor tyrosine-based activation motif. These findings were corroborated by analysis of tissue sections from virally infected macaques. Immunohistochemical staining revealed that Fas ligand expression was efficiently up-regulated in the GALT of animals that had been experimentally infected with wild-type SIV smmPBj14 but not in animals that were infected with a nonacutely pathogenic viral mutant lacking the Nef ITAM. Taken together, these results suggest that the ability of SIV smmPBj14 to cause acutely lethal disease and to up-regulate FasL expression may be linked. Additional studies will be required to determine whether the induction of FasL expression is in itself important for acute disease pathogenesis.

Journal of NeuroVirology, 1996

Quinolinic acid (Quin) is thought to underlie cognitive and motor dysfunctions for a variety of n... more Quinolinic acid (Quin) is thought to underlie cognitive and motor dysfunctions for a variety of neurological disorders. Specifically, in human immunodeficiency virus (HIV)-associated dementia, Quin levels correlate with the degree of neurological dysfunction observed in affected individuals. Since recent data from our laboratories suggest that both HIV-1 infection and activation of brain macrophages are required for the development of neurotoxicity we examined Quin production during virus infection and immune activation. HIV-1 infection of monocytes induced low levels of Quin while lipopolysaccharide (LPS) or interferon-gamma (IFN-gamma) activation of the virus-infected cells elicited 10-fold higher levels. The combined effects of LPS and IFN-gamma for Quin production in HIV-infected monocytes was identical to each factor added alone. Little or no Quin was detected in unstimulated uninfected monocytes. LPS or IFN-gamma activation of uninfected monocytes produced substantially higher levels of Quin than found in similarly stimulated HIV-1-infected monocytes. These results were at variance to the production of tumor necrosis factor-alpha (TNF-alpha). Here, a 2-to 5-fold increase in TNF-alpha levels were observed in culture fluids of LPS-activated HIV-infected cells when compared to similarly stimulated uninfected monocytes. The effect of LPS-induced Quin production by HIV-infected monocytes was not altered by primary human astrocytes. These data suggest that Quin levels seen in HIV dementia are a reflection of macrophage/ microglial activation seen during advanced clinical disease. These findings could help explain, in part, why few HIV-1-infected brain macrophages can give rise to significant neurological impairments.

BioTechniques, Jun 1, 1997

A method for simultaneously detecting membrane permeability (characteristic of necrosis) and DNA ... more A method for simultaneously detecting membrane permeability (characteristic of necrosis) and DNA fragmentation (characteristic of apoptosis) is described. By combining a common dye-exclusion method (Trypan Blue) with a commercially available terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end-labeling (TUNEL) labeling kit, we have succeeded in developing a novel methodology for obtaining permanently mounted slides of monolayer cell cultures double-labeled for DNA fragmentation and cell lysis. This method should facilitate in situ studies of cell death by allowing for a more accurate quantification of total toxicity in monolayer cell cultures and perhaps further enhance our understanding of the different mechanisms of cell death as well.

Journal of Neuroscience Research, Nov 15, 2004

Methods for growing primary neuronal cultures rely on the inclusion of antioxidants in the cultur... more Methods for growing primary neuronal cultures rely on the inclusion of antioxidants in the culture medium, but no studies have determined precisely if or when antioxidants are required for neuronal survival, despite the significance this information would have for understanding neurodevelopment and studying oxidative stress. We show that cortical neurons grown in Neurobasal media with B27 supplement required antioxidants for only the first 24 hr post‐explantation, after which the antioxidants could be removed permanently without noticeable loss of neuronal survival over the normal lifespan. Cortical cultures never exposed to antioxidants did not survive. These findings represent a novel method for substantially antioxidant‐free neuronal culture, whereby antioxidants can be removed permanently from the cultures after only 1 day. This method may prove critical for studies of oxidative stress, because B27 antioxidants significantly diminished pro‐oxidative effects of the excitatory neurotransmitter glutamate and hydrogen peroxide on cortical cultures, even if antioxidants were removed before the oxidizing treatment. Together, these findings suggest a brief window of high vulnerability to reactive oxygen species, and have important implications for studies of oxidative stress and developmental neuroscience. © 2004 Wiley‐Liss, Inc.

Virus Research, May 1, 1994

Recent studies in our laboratory and that of Dr. Howard Gendelman have revealed two important pat... more Recent studies in our laboratory and that of Dr. Howard Gendelman have revealed two important pathways for neuronal damage during HIV-l encephalopathy in children. First, substantial numbers of astrocytes are actively or latently infected with HIV-l. Astrocyte infection may lead to neuronal dysfunction through loss of supporting growth factors, excitotoxicity due to dysregulation of neurotransmitter reuptake, and loosening of the blood-brain barrier permitting further seeding of HIV-l in the CNS. Significantly, infection of astrocytes is marked by near-exclusive synthesis of early regulatory gene products of HIV-l, while structural proteins characteristic of productive infection are found in macrophages, microglia and multinucleated giant cells. We propose the term 'restricted' to denote the non-productive infection found in astrocytes. Second, HIV-l-infected macrophages initiate inflammatory processes which are amplified through cell-cell interactions with astrocytes. Macrophage-astrocyte interactions produce arachidonic metabolites and potentially neurotoxic cytokines 0'NF-a and IL-lfi), leading to astroglial activation and proliferation which then amplifies these cellular processes. These new findings suggest that two major pathways leading to neurotoxicity in pediatric AIDS encephalopathy are linked to HIV-l infection through astrocyte-mediated processes, and help explain how small numbers of productively infected cells indirectly cause widespread tissue pathology and elicit profound neurological impairment.

Neurology, Apr 22, 2012

Objective: To test whether URMC-099, a novel small molecule inhibitor of fms-like tyrosine kinase... more Objective: To test whether URMC-099, a novel small molecule inhibitor of fms-like tyrosine kinase 3 (FLT3) and mixed lineage kinase 3 (MLK3), can attenuate microglial activation and prevent neuronal injury in the hippocampus of mice with experimental autoimmune encephalomyelitis (EAE). Background Cognitive impairment in patients with multiple sclerosis (MS) correlates with widespread gray matter atrophy. Loss of hippocampal synapses, associated with microglial activation and demyelination in MS patients and an EAE mouse model, likely contributes to cognitive decline in MS but how this occurs and how to prevent it are unknown. We hypothesized that attenuation of microglial activation by kinase inhibitor URMC-099 would prevent synaptic loss and neuronal injury in the hippocampus of EAE mice. Design/Methods: We induced EAE in C57BL/6 mice via direct immunization with myelin oligodendrocyte glycoprotein peptide (MOG35-55, 100 µg) or control (saline) in complete Freund9s adjuvant with pertussis toxin (300 ng, days 0 and 2), and treated with vehicle or URMC-099 (50 mg/kg i.p. daily) beginning with the onset of clinical symptoms on day 10. After 4 weeks mice were perfused with paraformaldehyde, brains were sectioned and immunostained for myelin (MBP), monocyte/microglia (Iba1), presynaptic terminals (synapsin1) and dendrites (MAP2), and imaged by grid confocal fluorescence microscopy. Results: URMC-099 did not affect clinical EAE scores but reduced hippocampal monocyte/microglia activation to control levels (vs. 1.23x controls in vehicle-treated EAE mice, p Conclusions: Pharmacologic inhibition of FLT3 and MLK3 can prevent neuronal injury associated with microglial activation in EAE hippocampus, and may have potential for treatment of neurodegeneration and cognitive impairment in patients with MS. Supported by: NIH grants T32 NS 007338 and PO1 MH064570-10. Disclosure: Dr. Bellizzi has nothing to disclose. Dr. Gelbard has received personal compensation in an editorial capacity for the Journal of Neuroimmune Pharmacology.

Journal of Neuroinflammation, May 5, 2018

Background: The mixed lineage kinase type 3 inhibitor URMC-099 facilitates amyloid-beta (Aβ) clea... more Background: The mixed lineage kinase type 3 inhibitor URMC-099 facilitates amyloid-beta (Aβ) clearance and degradation in cultured murine microglia. One putative mechanism is an effect of URMC-099 on Aβ uptake and degradation. As URMC-099 promotes endolysosomal protein trafficking and reduces Aβ microglial pro-inflammatory activities, we assessed whether these responses affect Aβ pathobiogenesis. To this end, URMC-099's therapeutic potential, in Aβ precursor protein/presenilin-1 (APP/PS1) double-transgenic mice, was investigated in this model of Alzheimer's disease (AD). Methods: Four-month-old APP/PS1 mice were administered intraperitoneal URMC-099 injections at 10 mg/kg daily for 3 weeks. Brain tissues were examined by biochemical, molecular and immunohistochemical tests. Results: URMC-099 inhibited mitogen-activated protein kinase 3/4-mediated activation and attenuated βamyloidosis. Microglial nitric oxide synthase-2 and arginase-1 were co-localized with lysosomal-associated membrane protein 1 (Lamp1) and Aβ. Importatly, URMC-099 restored synaptic integrity and hippocampal neurogenesis in APP/PS1 mice. Conclusions: URMC-099 facilitates Aβ clearance in the brain of APP/PS1 mice. The multifaceted immune modulatory and neuroprotective roles of URMC-099 make it an attractive candidate for ameliorating the course of AD. This is buttressed by removal of pathologic Aβ species and restoration of the brain's microenvironment during disease. Highlights The therapeutic potential of the MLK3 inhibitor URMC-099 was evaluated in an AD mouse model. URMC-099 facilitates Aβ clearance and microglial morphological changes in diseased brain tissue. URMC-099 restores synaptic integrity and facilitates hippocampal neurogenesis in APP/PS1 mice. The multifaceted roles of URMC-099 make it an attractive therapeutic candidate for AD.

bioRxiv (Cold Spring Harbor Laboratory), Aug 31, 2023

Background: Links between acute lung injury (ALI), infectious disease, and neurological outcomes ... more Background: Links between acute lung injury (ALI), infectious disease, and neurological outcomes have been frequently discussed over the past few years, especially due to the COVID-19 pandemic. Yet, much of the cross-communication between organs, particularly the lung and the brain, has been understudied. Here, we have focused on the role of neutrophils in driving changes to the brain endothelium with ensuing microglial activation and neuronal loss in a model of ALI. Methods: We have applied a three-dose paradigm of 10μg/40μl intranasal lipopolysaccharide (LPS) to induce neutrophilia accompanied by proteinaceous exudate in bronchoalveolar lavage fluid (BALF) in adult C57BL/6 mice. Brain endothelial markers, microglial activation, and neuronal cytoarchitecture were evaluated 24hr after the last intranasal dose of LPS or saline. C57BL/6-Ly6g(tm2621(Cre-tdTomato)Arte (Catchup mice) were used to measure neutrophil and blood-brain barrier permeability following LPS exposure with intravit...

Ageing Research Reviews, Dec 1, 2021

Involvement of extracellular matrix (ECM) components in aging and age-related neurodegeneration i... more Involvement of extracellular matrix (ECM) components in aging and age-related neurodegeneration is not well understood. The role of hyaluronan (HA), a major extracellular matrix glycosaminoglycan, in malignancy and inflammation is gaining new understanding. In particular, the differential biological effects of high molecular weight (HMW-HA) and low molecular weight hyaluronan (LMW-HA), and the mechanism behind such differences are being uncovered. Tightly regulated in the brain, HA can have diverse effects on cellular development, growth and degeneration. In this review, we summarize the homeostasis and signaling of HA in healthy tissue, discuss its distribution and ontogeny in the central nervous system (CNS), summarize evidence for its involvement in age-related neurodegeneration and Alzheimer Disease (AD), and assess the potential of HA as a therapeutic target in the CNS.

Multiple sclerosis (MS) is an inflammatory, neurodegenerative disease of the CNS characterized by... more Multiple sclerosis (MS) is an inflammatory, neurodegenerative disease of the CNS characterized by both grey and white matter injury. Microglial activation and a reduction in synaptic density are key features of grey matter pathology that can be modeled with experimental autoimmune encephalomyelitis (EAE). Complement deposition combined with microglial engulfment has been shown during normal development and in disease as a mechanism for pruning synapses. We tested whether there is excess complement production in the EAE hippocampus and whether complement-dependent synapse loss is a source of degeneration in EAE using C1qa and C3 knockout mice. We found that C1q and C3 protein levels were elevated in EAE mice. Genetic loss of C1qa provided a small degree of protection from EAE-induced decreases in synaptic density. However, C1qa knockout EAE mice had similar levels of microglial activation and identical clinical scores as WT EAE mice. C3 knockout mice were largely protected from EAE-induced synapse loss and microglial activation, results that correlated with a reduction in the EAE clinical score. Thus, pathologic expression and activation of the early complement pathway drives a portion of the synapse elimination observed in the EAE grey matter.

Neuropathology and Applied Neurobiology, Jun 1, 1997

negative controls, had increased expression of Bcl-2 and Bcl-x in reactive astrocytes in cortex a... more negative controls, had increased expression of Bcl-2 and Bcl-x in reactive astrocytes in cortex and basal ganglia. In vitro studies using Western blot analysis demonstrated an up-regulation in the levels of Bax, and phosphorylated (i.e. inactive) Bcl-2 in HIV-1 infected macrophages, and in LPS-activated macrophages, relative to levels in virus-negative unstimulated macrophages. These results suggest that productive HIV-1 infection, or cellular activation, renders macrophages more vulnerable to apoptosis. Taken together, these findings suggest that brainresident macrophages and microglia in patients with HIV-1 encephalitis are more prone to undergo apoptosis and that astrocytes in contrast may be resistant to apoptosis. This may represent a mechanism to limit microglial activation and the spread of productive HIV-1 infection in the CNS of children with HIV-1 encephalitis.

Elsevier eBooks, 2004

This chapter reviews the epidemiology, clinical manifestations, and pathology of HIV-1-associated... more This chapter reviews the epidemiology, clinical manifestations, and pathology of HIV-1-associated neurological disease and its associated encephalitis. Particular emphasis is placed on diagnosis, pathogenesis, and treatment options now available or under investigation. This chapter demonstrates how HIV-1 infection of the brain results in neuronal injury and death. Besides the significant morbidity induced by HIV-1 for cognitive, motor, and behavioral function, the CNS represents an important reservoir for continuous virus production and the emergence of viral strain variance. Moreover, the mechanisms of how mononuclear phagocytes (MP) secretory products may damage neurons have potential wide applicability for other neurodegenerative disorders where inflammation is an important component of disease pathogenesis. Adjunctive therapies such as N-methyl D-aspartate receptor (NMDAR) blockers, chemokine and cytokine effectors, antioxidants, caspase inhibitors, and p38MAPK inhibitors are all potential interventions and are being developed by many laboratories. If successful, these drugs may find utility in a broad range of neurodestructive processes.

Journal of Neuroimmune Pharmacology, Feb 27, 2007

The phospholipid mediator platelet-activating factor (PAF), an endogenous modulator of glutamater... more The phospholipid mediator platelet-activating factor (PAF), an endogenous modulator of glutamatergic neurotransmission, can also be secreted by brain mononuclear phagocytes during HIV-1 infection. Platelet-activating factor can induce neuronal apoptosis by NMDA receptor-dependent and independent mechanisms. We now demonstrate that acute administration of sublethal doses of PAF to striatal slices augments synaptic facilitation in striatal neurons following high-frequency stimulation, which can be blocked by PAF receptor antagonists, suggesting that striatal synaptic facilitation can be augmented by PAF receptor agonism. We also demonstrate that repeated sublethal doses of PAF during tetanic stimulation can greatly increase the magnitude of postsynaptic potentials and action potentials, but a lethal dose of PAF destroys the capacity of corticostriatal synapses to achieve this augmented synaptic facilitation. Thus, the relative concentration and temporal pattern of PAF expression at glutamatergic synapses may govern whether it acts in a physiologic or pathophysiologic manner during striatal neurotransmission.

Neuron, Nov 1, 2020

Mordes et al. (2020) did not detect the survival or motor phenotypes in C9orf72 BAC transgenic mi... more Mordes et al. (2020) did not detect the survival or motor phenotypes in C9orf72 BAC transgenic mice originally described by Liu et al. (2016). We discuss methodological differences between the Mordes and Liu studies, several additional studies in which survival and motor phenotypes were found, and possible environmental and genetic effects. First, Nguyen et al. (2020) showed robust ALS/FTD phenotypes in C9-BAC versus non-transgenic (NT) mice and that α-GA1 treatment improved survival, behavior, and neurodegeneration. The groups of Gelbard and Saxena also show decreased survival of C9-BAC versus NT mice and neuropathological and behavioral deficits similar to those shown by Liu et al. (2016). Although FVB/N mice can have seizures, increases in seizure severity and death of C9 and NT animals, which may mask C9 disease phenotypes, have been observed in recent C9-500 FVB/NJ-bred cohorts. In summary, we provide an update on phenotypes seen in FVB C9-BAC mice and additional details to successfully use this model. This Matters Arising Response paper addresses the Mordes et al. (2020) Matters Arising paper, published concurrently in Neuron.

Epilepsy Research, 1994

Amygdala kindling resulted in significant increases in the expression of D2 receptor mRNA in the ... more Amygdala kindling resulted in significant increases in the expression of D2 receptor mRNA in the nucleus accumbens and striatum 30 days following the last kindling stimulation. Densitometric analyses of tissue sections incubated in the presence of an oligonucleotide probe directed against Dz receptor cDNA indicated a 2&35% increase in D2 receptor mRNA in these regions following kindling. Kindling from the amygdala followed by piriform cortical kindling in the transfer paradigm (overkindling) resulted in significant further increases in D2 receptor mRNA expression in both the accumbens (150% increase) and striatum (120% increase). There were no observed hemispheric asymmetries in D2 receptor mRNA in either kindled or overkindled animals. The data indicate an enduring upregulation of extrapyramidal Dz receptor mRNA following the kindling process. How this change may relate to kindling-induced alterations in seizure susceptibility or behaviors mediated by limbic dopaminergic pathways are questions for future studies.

Neuropathology and Applied Neurobiology, Oct 1, 1999

Apoptosis of neurones, macrophages, and microglia occurs in the brains of paediatric patients wit... more Apoptosis of neurones, macrophages, and microglia occurs in the brains of paediatric patients with human immunodeficiency virus (HIV) type 1 encephalitis, which is often associated with pre‐mortem neurological disease (progressive encephalopathy). We have previously reported that TUNEL‐positive neurones in brain tissue from paediatric patients with HIV type 1 encephalitis and progressive encephalopathy are strikingly devoid of the pro‐apoptotic gene product Bax, in marked contrast to brain‐resident macrophages and microglia. Using immunocytochemical methods, the present study demonstrate that neurones in patients with HIV type 1 encephalitis and progressive encephalopathy, as well as macrophages and microglia, but not astrocytes, overexpress caspase‐3, a pro‐apoptotic enzyme that is proteolytically activated downstream of Bax–Bcl‐2 dysregulation. Co‐localization of neuronal cytoplasmic caspase‐3 and nuclear TUNEL staining, a marker for fragmented DNA, was also infrequently observed in brain tissue from patients with HIV type 1 encephalitis and progressive encephalopathy. These findings suggest that vulnerable neurones in brain tissue from patients with HIV virus type 1 encephalitis and progressive encephalopathy undergo apoptosis by a mechanism that involves upregulation of caspase‐3 in a pathway that is independent of Bax–Bcl‐2 dysregulation. Furthermore, caspase‐3 upregulation in apoptotic neurones likely occurs prior to DNA fragmentation.

Elsevier eBooks, 2009

Diminution in synaptic function and loss of normal synaptic architecture are likely to be the bio... more Diminution in synaptic function and loss of normal synaptic architecture are likely to be the biologically relevant substrate for neurologic deficits in patients with HIV-1-associated dementia (HAD) and patients suffering from the prionoses. Because synaptic injury frequently precedes neuronal apoptosis and surviving neurons retain a remarkable capacity for synaptic repair and functional recovery, synaptic protection with neuroprotective agents may have great potential to improve neurologic function in HAD. In this article we focus on the evidence to support these claims, as well as discuss findings that explain how disruption of synaptic structure and function that can occur during excitotoxic injury with neuroinflammation represents a likely substrate for neurologic deficits. Based on available data, we provide a rationale for identification of molecular targets for synaptic protection in neurodegenerative disease that accompanies HIV-1 infection of the nervous system. While patients with HAD beginning antiretroviral therapy have shown reversal of neurologic symptoms that is unique for patients with chronic neurodegenerative conditions, we propose that the therapeutic potential for such reversal may not be limited to this disease.